JCI table of contents, February 2, 2004

01/26/04

1. Understanding the autoimmune response in type 1 diabetes

Type 1 diabetes mellitus (T1DM) is the result of immune-mediated destruction of insulin-secreting pancreatic beta cells. For more than 25 years researchers have searched for an environmental agent or event that triggers this autoimmune response. Past research has suggested that T cells that react to islet beta cells can contribute to the autoimmune response in diabetic patients and also play a part in self-tolerance in healthy individuals. The rarity of these cells and inadequate technology has impaired the examination of this paradigm. In the February 2 issue of the Journal of Clinical Investigation Mark Peakman and colleagues from King's College London suggest a mechanism for the specificity of this immune regulation that explains why the same peptides present on pancreatic b cells that activate T cells in patients with T1DM and normal individuals cause an autoimmune response in diabetic patients, but no such response in normal individuals.

The authors developed a novel assay to examine T cell responses to a panel of epitopes naturally expressed by islet cells and demonstrated that it is the pathways of T cell differentiation and maturation in reaction to these epitopes in T1DM patients (in whom autoimmunity develops) and normal individuals (in whom autoimmunity is arrested) that are different. Upon exposure to antigen, naïve T cells in normal individuals differentiate into T cells that produce IL-10, and possibly TGF- beta, subsequently inhibiting cells that would normally mediate an aggressive immune response. The results reported by Peakman and colleagues suggest that in patients with T1DM, there is instead induction of a predominant number of T cells that produce IFN-gamma and IL-2, which drives an autoaggressive immune response. Why these T cell activation pathways differ between normal and T1DM patients will require further characterization.

In an accompanying commentary, Kevan Herald from the Naomi Berrie Diabetes Center at Columbia University, New York, comments that "these new findings concerning the responses of normal individuals and patients with T1DM to autoantigens shed light on the differences in immune responses between these two groups and the mechanisms of pathogenesis of the disease. The findings suggest ways in which regulation of the autoimmune response occur and offer approaches to immune modulation and ultimately tolerance, the immunologist's "Holy Grail."

TITLE: Autoreactive T cell responses show proinflammatory polarization in diabetes but a regulatory phenotype in health

The outermost layer of the skin – the epidermis – is a rapidly renewing tissue and relies on the regenerative capacity of keratinocytes. Skin grafts using human cultured epidermal cells have been successful in treating patients with severe skin wounds. The notion that the ability to regenerate functional epidermal tissue is an exclusive property of epidermal stem cells is a general assumption in the stem cell biology field. In the February 2 issue of the Journal of Clinical Investigation, Pritinder Kaur and colleagues at the Peter MacCallum Cancer Centre, Australia, demonstrate that both epidermal stems cells and their early, differentiated progeny contribute to rapid epidermal regeneration.

The majority of proliferating epidermal cells, also known as transit-amplifying cells, at the inner-most layer of the skin have a finite life span and undergo rapid terminal differentiation. Therefore it is well accepted that the extensive regenerative capacity of the skin is most likely attributed to the activity of epidermal stem cells.

To determine the cells responsible for rapid epidermal regeneration, Kaur and colleagues separated epidermal stem cells from their progeny and assayed the ability of both cell types to regenerate epidermal tissue in both in vitro and in vivo settings. As expected, keratinocyte stem cells displayed robust regenerative capabilities, but unexpectedly, transit-amplifying cells and early differentiating cells, which are more committed progenitor cells, could also form a fully stratified epidermis under appropriate microenvironmental conditions. The authors also demonstrated that the regenerative capacity of these cell types could be enhanced by exposure to the protein laminin-10/11.

This work presents important new considerations for the expansion of keratinocyte progenitor cell populations for therapies that require large numbers of epidermal cells, such as those required for the treatment of severe wounds such as extensive burns. It may be possible to harness the vast proliferative potential of readily available and accessible keratinocyte progenitors of the skin for cellular therapies, thereby removing the need for difficult and limited stem cell selection.

Antitumor immunotherapy is a challenging endeavor since most human tumor–associated antigens are nonmutated self-proteins expressed on normal tissues. The ideal vaccination approach requires self-tolerance while eliciting an effective antitumor response. In an effort to design the ideal vaccine, David-Alexandre Gross and colleagues from Institut Gustave Roussy, France, examined epitopes (parts of antigen molecules) derived from murine telomerase reverse transcriptase (mTERT) – an enzyme expressed in more than 80% of human tumors – as potential anticancer vaccines. In the February 2 issue of the Journal of Clinical Investigation they report that epitopes with a low affinity for HLA class I molecules, molecules which bind and present antigens to T cells for killing – could be modified to generate an immune response against the tumor without stimulating an immune response against self. Mice vaccinated with the modified low-affinity epitopes maintained tumor immunity, while mice vaccinated with high-affinity epitopes died after challenge with tumor cells. These studies highlight the importance of rational epitope selection for effective cancer vaccines.

TITLE: High vaccination efficiency of low-affinity epitopes in antitumor immunotherapy

Organic nitrates, such as nitroglycerin, are widely used in the treatment of coronary artery disease and congestive heart failure. Despite clear benefits when given acutely, organic nitrates lose their clinical effectiveness when used as sustained therapy – a phenomenon known as nitrate tolerance. In the February 2 issue of the Journal of Clinical Investigation. Thomas Münzel and colleagues from The University Hospital Eppendorf, Germany, demonstrate that it is the dysfunction of the enzyme mitochondrial aldehyde dehydrogenase, responsible for denitrification of organic nitrates, that causes nitrate tolerance.

In an accompanying commentary, John D. Parker from Mount Sinai Hospital, Canada, comments "the concept that sustained nitrate therapy causes increased production of free radicals has a number of important implications. Although their clinical efficacy during chronic therapy had been questioned for many years because of phenomena of tolerance, it had generally been assumed that the organic nitrates were safe drugs with no potential for long-term toxicity. The present study, remarkably documents that therapy with nitroglycerin causes increased free radical production by cardiac mitochondria. These observations suggest that the time has come to mount an appropriately powered clinical trial examining the efficacy and safety of sustained therapy with organic nitrates for those diseases for which they are commonly used."

TITLE: Central role of mitochondrial aldehyde dehydrogenase and reactive oxygen species
in nitroglycerin tolerance and cross-tolerance

Maple syrup urine disease (MSUD) is an inherited disorder caused by deficiency of the enzyme branched-chain alpha-keto acid dehydrogenase involved in amino acid metabolism, and is characterized by urine that smells like maple syrup. In the February 2 issue of the Journal of Clinical Investigation, Yuan-Tsong Chen and colleagues from Academia Sinica, Taiwan, screened families of mutant mice for disorders of amino acid metabolism using tandem mass spectrometry (MS/MS). This technique is now the choice for newborn screening for up to 40 metabolic disorders within both the USA and Europe. Through the use of mutagenesis in mice and screening of their offspring using MS/MS, a novel disorder similar to MSUD was discovered. The authors found elevated levels of branched-chain amino acids, suggesting a disorder similar to MSUD, however they report a mutation in another enzyme in this pathway – branched-chain aminotransferase. Chen et al. then utilized their newly found mouse model of MSUD to assess the dietary management of MSUD and found that the muscular weakness, hair loss, and premature death observed in mice with MSUD, could be ameliorated or prevented by a diet low in branched-chain amino acids.

In an accompanying commentary, Arnold Strauss from Vanderbilt Children's Hospital, Tennessee, comments "because mouse and human metabolism is similar, it is very likely that humans with a similar clinical and biochemical phenotype will turn out to have mutations in this gene." He continues "this result demonstrates the important general principle that murine mammalian models of human disorders can be used to determine the natural history of such disorders and quickly assess outcomes of treatment, studies that are often impossible to complete in humans and, even if feasible, take years to perform." A widespread application of the mutagenesis approach reported by Chen and co-workers should facilitate the identification and characterization of genetically modified disorders directly relevant to human disease.

Lamin A and C are proteins that form a thin fibrillar web between the nuclear matrix and nuclear envelope, which helps maintain nuclear structure and function. Mutations in the LMNA gene, which encodes these proteins, cause a variety of diseases affecting striated muscle, fat cells, and nerves. It has been unclear how mutations in these proteins, expressed in many different cell types, cause different diseases. Two independent studies in the February 2 issue of the Journal of Clinical Investigation examine lamin A/C–deficient mice and report that, during mechanical stress, lamin A/C–deficient cells develop abnormalities in their distribution of chromatin and nuclear envelope damage, which results in secondary alterations in the genes normally activated in adaptive and protective responses to stress.

Diane Fatkin and colleagues from the Victor Change Cardiac Research Institute in Australia demonstrated that lamin A/C–deficient mice develop dilated cardiomyopathy. The nuclei within cardiac cells possessed abnormal nuclear structure that possibly contributes to defects in the transport of molecules in and out of the nucleus, as well as alterations in gene expression resulting in cardiomopathy.

In the second study, Richard Lee and colleagues from the Massachusetts Institute of Technology found that fibroblasts from lamin A/C–deficient mice had misshaped nuclei, structural damage, and decreased stiffness that was increased in response to increased mechanical stress. They also observed alterations in gene expression.

In an accompanying commentary, Howard Worman and Jean-Claude Courvalin from Columbia University and Université Paris, respectively, discuss how impaired nuclear structure and altered gene expression may cause damage in a diversity of tissue types.

Antibiotic exposure not the only cause of antibiotic resistance in cystic fibrosis

The organism Burkholderia cepacia is associated with severe morbidity and mortality in patients with cystic fibrosis, and is resistant to many common antibiotics. This resistance is mediated by the inability of some antibiotics to bind to the outer membrane of the bacterium, or antibiotic-degrading enzymes that the organism releases. In the February 2 issue of the Journal of Clinical Investigation, Jane Burns and colleagues from the Children's Hospital and Regional Medical Center in Seattle identify a cluster of genes that encode an antibiotic efflux pump, which facilitates the active removal of a variety of antibiotics from the organism so that any intracellular antibiotic concentrations are too low to be effective at the target site. Efflux was enhanced in the presence of salicylate and low iron concentratrions suggesting that salicylate is a natural substrate for the pump and that the environment of low iron concentrations in the cystic fibrosis lung can induce efflux-mediated antibiotic resistance, even in the absence of the antibiotic selective pressure that drives selection of antibiotic-resistant bacterial mutants.

The report indicates that induction of antibiotic resistance in the host may not be solely determined by antibiotic exposure, a finding that has the potential to impact physicians' management of cystic fibrosis airway infections.

Leptin aims for the central nervous system in the control of defective fat metabolism

Leptin administration has been highly successful in treating the diabetes, insulin resistance, and fatty liver degeneration associated with lipodystrophy – a disturbance in fat metabolism. This hormone, expressed by fat cells, acts on a variety of tissues including the brain, skeletal muscles, heart, and pancreatic beta cells. In the February 2 issue of the Journal of Clinical Investigation, Jeffrey Friedman and colleagues from The Rockefeller University in New York have identified the central nervous system as the primary site of action for leptin's effects on metabolic improvement in lipodystrophy.

Using a congenital mouse model of lipodystrophy, the authors observed that administration of leptin under the skin was not as potent as lower doses administered directly to the central nervous system, which were able to correct the metabolic abnormalities associated with lipodystrophy. Furthermore, using microarray technology, the authors identified repression of the enzyme SCD-1 as a mechanism of leptin action to improve fatty liver degeneration. These findings may have profound effects on the progression of leptin treatment strategies for metabolic diseases.

TITLE: Site and mechanism of leptin action in a rodent form of congenital lipodystrophy

The activation of platelets contributes to the formation of thrombi in the circulation, which block arteries and can cause heart attack and stroke. Interested in the role of the serine/threonine kinase, Akt, in platelet activation, Donna Woulfe and colleagues from the University of Pennsylvania examined platelets from normal and Akt2-deficient mice. In the February 2 issue of the Journal of Clinical Investigation the authors report that platelets from Akt2-deficient mice, but not Akt1-deficient mice, have defects in aggregation, secretion, and thrombus formation. Furthermore, Akt2 was shown to be more highly expressed in mouse platelets than Akt1. The data indicate that Akt is
needed for optimal platelet aggregation, fibrinogen binding, and secretion which is critical for stabilizing thrombus formation after arterial injury.

Enzyme IKK2 found to play no major role in obesity-induced insulin resistance

The enzyme IkappaB kinase, IKK2, plays a major role in regulating immune and inflammatory responses. In the February 2 issue of the Journal of Clinical Investigation, Jens Brüning and colleagues from the University of Cologne, Germany, investigate the role of IKK2 in obesity-induced insulin resistance. Using mice with a muscle-specific knockout of IKK2 as well as mice exhibiting a 50% reduction of IKK2 expression in every tissue, the authors report that both normal and mutant mice developed obesity to a similar extent. The data argue against a substantial role for muscular IKK2 in mediating obesity-induced insulin resistance.

TNF-alpha is a cytokine produced by innate immune and Th1 cells known to be involved in protective (type 1) immunity against bacterial infection. Zhou Xing and colleagues from McMaster University, Canada, and Institut Pasteur, France, sought to elucidate the role of this cytokine in the type 1 immune response by characterizing TNF-alpha–deficient mice upon challenge with Mycobacterium bovis BCG. In the February 2 issue of the Journal of Clinical Investigation the authors report that mice deficient in TNF-alpha died upon pulmonary infection with myobacteria and suffered from an overactivation of the type 1 immune response as evidenced by expansion of CD4 and CD8 cells, increased frequency of antigen-specific T cells, overproduction of proimmune cytokines IFN-gamma and IL-12, and severe lung injury. Establishing TNF-alpha as a negative regulator of type 1 immunity has important implications for the development of therapeutic strategies that aim to block TNF-alpha.

Glycoprotein 130: star of one signaling pathway with multiple outcomes for healthy bones

Mammalian skeletal growth and maintenance involves cooperation among bone formation, breakdown, and remodeling. Cytokines such as IL-6, dependent on glycoprotein 130 (gp130), are known to signal through at least 2 intracellular pathways: STAT1/3 and SHP2/ras/MAPK. However, the distinct downstream effects of these two pathways on bone development were previously unknown.

In the February 2 issue of the Journal of Clinical Investigation. Natalie Sims and colleagues from The University of Melbourne, Australia, resolve the effects of these signaling pathways by targeted mutagenesis of gp130 in mice, which rendered them deficient in either STAT1/3 activation or SHP2/ras/MAPK activation. The authors report that STAT1/3 activation is involved in the proliferation of chondrocytes and osteoblasts – cells crucial to bone formation, whereas SHP2/ras/MAPK activation plays a key role in inhibiting osteoclasts – cells causing bone breakdown. These insights may provide new therapeutic targets for improving bone growth and bone density in mammals.